RNA regulation provides a mechanism to rapidly control gene expression in response to stimuli, including environmental changes. This project seeks to generate and utilize structural information to enhance our understanding of these processes with an emphasis on the importance of RNA target specificity for proper gene regulation. In this fiscal year, we have studied the atomic structures and functions of different RNA-binding proteins that regulate germline development, cellular responses to the environment, and ribosomal biogenesis. A major focus of our group is to understand the function of Pumilio/FBF (PUF) proteins. We determined the first crystal structure of a PUF protein in complex with RNA, which allowed us to understand the RNA recognition properties of PUF proteins and have extended these structural studies to a variety of PUF proteins. We have determined crystal structures of engineered proteins with altered RNA recognition specificity, and these results are described in a recently submitted manuscript. We have also determined the first crystal structures of a PUF protein and discovered a novel dimeric RNA recognition mechanism. We have also used crystal structures and biophysical and biochemical assays to study yeast Nop9 protein, an essential protein in small ribosomal subunit biogenesis that represents a second sub-family of PUF-like proteins. This new PUF-like protein has different RNA recognition properties than the classical PUF proteins. Our new studies follow up on our work published previously in Nature Communications.